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1.
Ecotoxicol Environ Saf ; 201: 110806, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32512418

RESUMO

The present study investigated the expressions of signalling molecules and inflammatory cytokines involved in copper-induced inflammatory responses of the mouse liver. A total of 240 institute of cancer research (ICR) mice (half male and half female) aged four weeks were randomly allocated to four groups treated with 0, 4, 8, and 16 mg/kg of [Cu] (Cu2+-CuSO4) for 42 days, respectively. [Cu] exceeding 4 mg/kg was found to induce inflammatory responses of the liver. Results showed significant up-regulation of mRNA and protein levels of apoptosis signal-regulating kinase 1 (ASK1), mitogen-activated protein kinase kinases 3/6 (MEK3/6), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38 MAPK), mitogen-activated protein kinase kinases 4/7 (MEK4/7), mitogen-activated protein kinase kinases 1/2 (MEK1/2), and extracellular signal-regulated protein kinases 1/2 (Erk1/2) due to Cu. By doing so, copper could activate the mitogen-activated protein kinases (MAPKs) signalling pathway. Concurrently, the nuclear factor-kappa B (NF-κB) signalling pathway was also activated in the Cu-treatment, as demonstrated by higher expressions of NF-κB and cyclooxygenase-2 (COX-2), activities of inducible nitric oxide synthase (iNOS), contents of nitric oxide (NO) and prostaglandin E2 (PGE2), and reducing levels of expression of inhibitory kappa B (IκB). High Cu intake also up-regulated expression levels of some pro-inflammatory mediators such as interleukin-2 (IL-2), interleukin-1ß (IL-1ß), and interleukin-8 (IL-8), and down-regulated the levels of expression of transforming growth factor beta (TGF-ß), an anti-inflammatory mediator. Additionally, our results indicated that Cu caused hepatic dysfunction, with evidence of occurrence of histopathological lesions and higher serum activities of alkaline phosphatase (AKP), aspartic acid transferase (AST), alanine amino transferase (ALT), and gamma-glutamyl transpeptidase (GGT), contents of albumin (ALB) and total bilirubin (TBIL). Altogether, the aforementioned results indicate that [Cu], at more than 4 mg/kg, induces the inflammatory responses in the liver via NF-κB and MAPKs signalling pathways, subsequently inducing hepatic dysfunction.


Assuntos
Cobre/toxicidade , Interleucina-1beta/metabolismo , Interleucina-2/metabolismo , Fígado/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Animais , Ciclo-Oxigenase 2/metabolismo , Feminino , Fígado/imunologia , Fígado/metabolismo , Fígado/patologia , Testes de Função Hepática , Masculino , Camundongos , Camundongos Endogâmicos ICR , Distribuição Aleatória , Transdução de Sinais
2.
Acta Cytol ; 64(5): 442-451, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32599588

RESUMO

INTRODUCTION: Persistent infection with high-risk human papillomavirus (HPV) types is associated with high-grade intraepithelial lesions (HSILs) and invasive cervical cancer. The host immune response plays a key role in whether HPV clears or persists. Most studies on local immune response to HPV collect cervical mucus in order to quantify secreted cytokines; however, cells located inside the tissue can release different cytokines associated with HPV infection. OBJECTIVE: This study compared the cytokine levels in cervical biopsy specimens of women with abnormal colposcopic findings according to the histopathological results: low-grade intraepithelial lesion (LSIL), HSIL, and no intraepithelial lesion (NSIL). METHODS: A cross-sectional study enrolling 141 cervical biopsy specimens examined the cytokine profile for interleukin (IL-) 2, IL-4, IL-10, IL-12, IL-17, and IL-23 and interferon-γ, using the Luminex assay/ELISA. Differences in cytokine levels among the cervical lesion groups were assessed using the Kruskal-Wallis test. RESULTS: The 141 specimens included 90 HSILs, 22 LSILs, and 29 NSILs. IL-2 levels were significantly higher in NSIL samples than in LSIL or in HSIL samples (p = 0.0001) and IL-23 levels were significantly higher in NSIL than in HSIL samples (p = 0.003). CONCLUSIONS: Our study shows that in samples from the lesion site point, 2 important pro-inflammatory cytokines, IL-2 and IL-23, are downregulated in HPV lesions.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasia Intraepitelial Cervical/patologia , Interleucina-23/metabolismo , Interleucina-2/metabolismo , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/complicações , Neoplasias do Colo do Útero/patologia , Adulto , Neoplasia Intraepitelial Cervical/metabolismo , Neoplasia Intraepitelial Cervical/virologia , Estudos Transversais , Regulação para Baixo , Feminino , Humanos , Infecções por Papillomavirus/metabolismo , Infecções por Papillomavirus/virologia , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/virologia
3.
Nat Commun ; 11(1): 2779, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32487999

RESUMO

T cell receptor (TCR) activation is modulated by mechanisms such as TCR endocytosis, which is thought to terminate TCR signalling. Here we show that, upon internalization, TCR continues to signal from a set of specialized endosomes that are crucial for T cell functions. Mechanistically, TCR ligation leads to clathrin-mediated internalization of the TCR-CD3ζ complex, while maintaining CD3ζ signalling, in endosomal vesicles that contain the insulin responsive aminopeptidase (IRAP) and the SNARE protein Syntaxin 6. Destabilization of this compartment through IRAP deletion enhances plasma membrane expression of the TCR-CD3ζ complex, yet compromises overall CD3ζ signalling; moreover, the integrity of this compartment is also crucial for T cell activation and survival after suboptimal TCR activation, as mice engineered with a T cell-specific deletion of IRAP fail to develop efficient polyclonal anti-tumour responses. Our results thus reveal a previously unappreciated function of IRAP-dependent endosomal TCR signalling in T cell activation.


Assuntos
Cistinil Aminopeptidase/metabolismo , Endossomos/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais/fisiologia , Linfócitos T/metabolismo , Animais , Membrana Celular/metabolismo , Proliferação de Células , Clatrina/metabolismo , Cistinil Aminopeptidase/genética , Modelos Animais de Doenças , Endocitose/fisiologia , Células HEK293 , Humanos , Interleucina-2/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Qa-SNARE/metabolismo , Transcriptoma
5.
Proc Natl Acad Sci U S A ; 117(11): 6047-6055, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32123114

RESUMO

Interleukin (IL)-2 and IL-21 dichotomously shape CD8+ T cell differentiation. IL-2 drives terminal differentiation, generating cells that are poorly effective against tumors, whereas IL-21 promotes stem cell memory T cells (TSCM) and antitumor responses. Here we investigated the role of metabolic programming in the developmental differences induced by these cytokines. IL-2 promoted effector-like metabolism and aerobic glycolysis, robustly inducing lactate dehydrogenase (LDH) and lactate production, whereas IL-21 maintained a metabolically quiescent state dependent on oxidative phosphorylation. LDH inhibition rewired IL-2-induced effects, promoting pyruvate entry into the tricarboxylic acid cycle and inhibiting terminal effector and exhaustion programs, including mRNA expression of members of the NR4A family of nuclear receptors, as well as Prdm1 and Xbp1 While deletion of Ldha prevented development of cells with antitumor effector function, transient LDH inhibition enhanced the generation of memory cells capable of triggering robust antitumor responses after adoptive transfer. LDH inhibition did not significantly affect IL-21-induced metabolism but caused major transcriptomic changes, including the suppression of IL-21-induced exhaustion markers LAG3, PD1, 2B4, and TIM3. LDH inhibition combined with IL-21 increased the formation of TSCM cells, resulting in more profound antitumor responses and prolonged host survival. These findings indicate a pivotal role for LDH in modulating cytokine-mediated T cell differentiation and underscore the therapeutic potential of transiently inhibiting LDH during adoptive T cell-based immunotherapy, with an unanticipated cooperative antitumor effect of LDH inhibition and IL-21.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Inibidores Enzimáticos/farmacologia , Interleucinas/metabolismo , L-Lactato Desidrogenase/antagonistas & inibidores , Melanoma Experimental/terapia , Células-Tronco/imunologia , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/transplante , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Linhagem Celular Tumoral/transplante , Humanos , Memória Imunológica , Imunoterapia Adotiva/métodos , Interleucina-2/imunologia , Interleucina-2/metabolismo , Interleucinas/imunologia , L-Lactato Desidrogenase/metabolismo , Melanoma Experimental/imunologia , Camundongos , Cultura Primária de Células , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo
6.
Proc Natl Acad Sci U S A ; 117(12): 6686-6696, 2020 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-32161126

RESUMO

Cytotoxic CD8+ T cells can effectively kill target cells by producing cytokines, chemokines, and granzymes. Expression of these effector molecules is however highly divergent, and tools that identify and preselect CD8+ T cells with a cytotoxic expression profile are lacking. Human CD8+ T cells can be divided into IFN-γ- and IL-2-producing cells. Unbiased transcriptomics and proteomics analysis on cytokine-producing fixed CD8+ T cells revealed that IL-2+ cells produce helper cytokines, and that IFN-γ+ cells produce cytotoxic molecules. IFN-γ+ T cells expressed the surface marker CD29 already prior to stimulation. CD29 also marked T cells with cytotoxic gene expression from different tissues in single-cell RNA-sequencing data. Notably, CD29+ T cells maintained the cytotoxic phenotype during cell culture, suggesting a stable phenotype. Preselecting CD29-expressing MART1 TCR-engineered T cells potentiated the killing of target cells. We therefore propose that CD29 expression can help evaluate and select for potent therapeutic T cell products.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Citotoxicidade Imunológica/imunologia , Integrina beta1/metabolismo , Interferon gama/metabolismo , Interleucina-2/metabolismo , Melanoma/patologia , Linfócitos T Citotóxicos/imunologia , Perfilação da Expressão Gênica , Humanos , Melanoma/imunologia , Melanoma/metabolismo , Prognóstico , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Taxa de Sobrevida
7.
Proc Natl Acad Sci U S A ; 117(13): 7183-7192, 2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32184322

RESUMO

Interleukin-2 (IL-2) is a small α-helical cytokine that regulates immune cell homeostasis through its recruitment to a high-affinity heterotrimeric receptor complex (IL-2Rα/IL-2Rß/γc). IL-2 has been shown to have therapeutic efficacy for immune diseases by preferentially expanding distinct T cell compartments, and several regulatory T cell (Treg)-biasing anti-IL-2 antibodies have been developed for combination therapies. The conformational plasticity of IL-2 plays an important role in its biological actions by modulating the strength of receptor and drug interactions. Through an NMR analysis of milliseconds-timescale dynamics of free mouse IL-2 (mIL-2), we identify a global transition to a sparse conformation which is regulated by an α-helical capping "switch" at the loop between the A and B helices (AB loop). Binding to either an anti-mouse IL-2 monoclonal antibody (mAb) or a small molecule inhibitor near the loop induces a measurable response at the core of the structure, while locking the switch to a single conformation through a designed point mutation leads to a global quenching of core dynamics accompanied by a pronounced effect in mAb binding. By elucidating key details of the long-range allosteric communication between the receptor binding surfaces and the core of the IL-2 structure, our results offer a direct blueprint for designing precision therapeutics targeting a continuum of conformational states.


Assuntos
Interleucina-2/metabolismo , Animais , Linhagem Celular , Interleucina-2/genética , Espectroscopia de Ressonância Magnética , Camundongos , Conformação Proteica
8.
Mol Biotechnol ; 62(5): 306-315, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32193710

RESUMO

NK cells have been seen as potential agents in adoptive immunotherapy for cancer. The main challenge for the success of this approach is to obtain a great quantity of activated NK cells for adoptive transfer. The present study had aimed to evaluate the effect of a feeder layer of irradiated MSCs in the in vitro expansion of NK cells. MSCs were obtained from the bone marrow (BM) cells remaining in the bag and filter used in the transplantation of hematopoietic stem cells. NK cells were obtained from peripheral blood (PB) of healthy volunteers. NK expansion and activation were stimulated by culture with artificial antigen-presenting cells (aAPCs) and IL-2, in the presence or absence of BM-MSCs. NK cell proliferation, phenotypic expression and cytotoxic activity were evaluated. Both culture conditions showed high NK purity with predominance of NK CD56brightCD16+ subset post expansion. However, cultures without the presence of MSCs showed higher NK proliferation, expression of activation markers (CD16 and NKG2D) and related cytotoxic activity. In this experimental study, the presence of a feeder layer of irradiated BM-MSCs interfered negatively in the expansion of PB-NKs, limiting their growth and activation. Further investigation is needed to understand the mechanisms of NK-MSC interaction and its implications.


Assuntos
Células Apresentadoras de Antígenos/citologia , Células Matadoras Naturais/citologia , Leucócitos Mononucleares/citologia , Células-Tronco Mesenquimais/citologia , Células Apresentadoras de Antígenos/imunologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Proteínas Ligadas por GPI/metabolismo , Regulação da Expressão Gênica , Voluntários Saudáveis , Humanos , Interleucina-2/metabolismo , Células K562 , Células Matadoras Naturais/imunologia , Leucócitos Mononucleares/imunologia , Células-Tronco Mesenquimais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Receptores de IgG/metabolismo
9.
J Appl Microbiol ; 129(3): 753-767, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32145053

RESUMO

AIMS: To examine the modulation of the interacting partners of the calcineurin (CaN)-NFAT pathway in T cells during Cryptococcus neoformans fungal infection and post-T11TS immunotherapy. METHODS AND RESULTS: Wistar rats were infected with C. neoformans and followed by immunotherapy with immune-potentiator T11TS. T cells were analysed by flow cytometry, immunoblotting and nuclear translocation study. The signalling proteins LCK, FYN, LAT, PLCγ1 and CaN in T cells were regulated by C. neoformans infection resulting in reduced nuclear translocation of NFAT and IL-2 expression. Following T11TS immunotherapy, the expressions of the above-mentioned proteins were boosted and thus resulting in the clearance of C. neoformans from lung and spleen. CONCLUSIONS: The precise mechanism of suppression of the T-cell function by C. neoformans is still unknown. Previously, we have shown that T11TS positively regulates the function of T cells to abrogate glioma and other immunosuppressive conditions. T11TS immunotherapy increased the expression of the above signalling partners of the CaN-NFAT pathway in T cells and improved nuclear retention of NFAT. As a result, an increased IL-2 expression leads to activation and proliferation of T cells. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results demonstrate the role of T11TS in restoring the CaN-NFAT signalling pathway in T cells. It identifies T11TS as an immunotherapeutic agent with potential clinical outcomes to counteract C. neoformans infection.


Assuntos
Antígenos CD58/uso terapêutico , Calcineurina/metabolismo , Criptococose/terapia , Fatores de Transcrição NFATC/metabolismo , Linfócitos T/imunologia , Animais , Criptococose/imunologia , Criptococose/microbiologia , Cryptococcus neoformans/efeitos dos fármacos , Cryptococcus neoformans/imunologia , Imunoterapia/métodos , Interleucina-2/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Ratos , Ratos Wistar , Ovinos , Transdução de Sinais/efeitos dos fármacos , Linfócitos T/metabolismo
10.
BMC Infect Dis ; 20(1): 185, 2020 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-32111171

RESUMO

BACKGROUND: Staphylococcus aureus (S. aureus) arthritis is one of the most detrimental joint diseases known and leads to severe joint destruction within days. We hypothesized that the provision of auxiliary immunoregulation via an expanded compartment of T regulatory cells (Tregs) could dampen detrimental aspects of the host immune response whilst preserving its protective nature. Administration of low-dose interleukin 2 (IL2) preferentially expands Tregs, and is being studied as a treatment choice in several autoimmune conditions. We aimed to evaluate the role of IL2 and Tregs in septic arthritis using a well-established mouse model of haematogenously spred S. aureus arthritis. METHODS: C57BL/6 or NMRI mice we intravenously (iv) injected with a defined dose of S. aureus LS-1 or Newman and the role of IL2 and Tregs were assessed by the following approaches: IL2 was endogenously delivered by intraperitoneal injection of a recombinant adeno-associated virus vector (rAAV) before iv S. aureus inoculation; Tregs were depleted before and during S. aureus arthritis using antiCD25 antibodies; Tregs were adoptively transferred before induction of S. aureus arthritis and finally, recombinant IL2 was used as a treatment starting day 3 after S. aureus injection. Studied outcomes included survival, weight change, bacterial clearance, and joint damage. RESULTS: Expansion of Tregs induced by IL2 gene therapy prior to disease onset does not compromise host resistance to S. aureus infection, as the increased proportions of Tregs reduced the arthritis severity as well as the systemic inflammatory response, while simultaneously preserving the host's ability to clear the infection. CONCLUSIONS: Pre-treatment with IL2 gene therapy dampens detrimental immune responses but preserves appropriate host defense, which alleviates S. aureus septic arthritis in a mouse model.


Assuntos
Artrite Infecciosa/prevenção & controle , Terapia Genética , Interleucina-2/genética , Staphylococcus aureus/patogenicidade , Animais , Anticorpos Monoclonais/uso terapêutico , Artrite Infecciosa/etiologia , Dependovirus/genética , Modelos Animais de Doenças , Feminino , Vetores Genéticos/metabolismo , Interleucina-2/metabolismo , Subunidade alfa de Receptor de Interleucina-2/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/uso terapêutico , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo
11.
Immunity ; 52(2): 313-327.e7, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-32049052

RESUMO

T cell responses upon infection display a remarkably reproducible pattern of expansion, contraction, and memory formation. If the robustness of this pattern builds entirely on signals derived from other cell types or if activated T cells themselves contribute to the orchestration of these population dynamics-akin to bacterial quorum regulation-is unclear. Here, we examined this question using time-lapse microscopy, genetic perturbation, bioinformatic predictions, and mathematical modeling. We found that ICAM-1-mediated cell clustering enabled CD8+ T cells to collectively regulate the balance between proliferation and apoptosis. Mechanistically, T cell expressed CD80 and CD86 interacted with the receptors CD28 and CTLA-4 on neighboring T cells; these interactions fed two nested antagonistic feedback circuits that regulated interleukin 2 production in a manner dependent on T cell density as confirmed by in vivo modulation of this network. Thus, CD8+ T cell-population-intrinsic mechanisms regulate cellular behavior, thereby promoting robustness of population dynamics.


Assuntos
Antígenos CD28/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/metabolismo , Animais , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Comunicação Celular , Contagem de Células , Linhagem Celular , Sobrevivência Celular , Rastreamento de Células , Células Dendríticas/imunologia , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-2/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Teóricos
12.
Sci Rep ; 10(1): 2886, 2020 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-32076052

RESUMO

A continuing quest for specific inhibitors of proinflammatory cytokines brings promise for effective therapies designed for inflammatory and autoimmune disorders. Cefazolin, a safe, first-generation cephalosporin antibiotic, has been recently shown to specifically interact with interleukin 15 (IL-15) receptor subunit α (IL-15Rα) and to inhibit IL-15-dependent TNF-α and IL-17 synthesis. The aim of this study was to elucidate cefazolin activity against IL-2, IL-4, IL-15 and IL-21, i.e. four cytokines sharing the common cytokine receptor γ chain (γc). In silico, molecular docking unveiled two potential cefazolin binding sites within the IL-2/IL-15Rß subunit and two within the γc subunit. In vitro, cefazolin decreased proliferation of PBMC (peripheral blood mononuclear cells) following IL-2, IL-4 and IL-15 stimulation, reduced production of IFN-γ, IL-17 and TNF-α in IL-2- and IL-15-treated PBMC and in IL-15 stimulated natural killer (NK) cells, attenuated IL-4-dependent expression of CD11c in monocyte-derived dendritic cells and suppressed phosphorylation of JAK3 in response to IL-2 and IL-15 in PBMC, to IL-4 in TF-1 (erythroleukemic cell line) and to IL-21 in NK-92 (NK cell line). The results of the study suggest that cefazolin may exert inhibitory activity against all of the γc receptor-dependent cytokines, i.e. IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21.


Assuntos
Anti-Inflamatórios/farmacologia , Cefazolina/farmacologia , Subunidade gama Comum de Receptores de Interleucina/antagonistas & inibidores , Adulto , Anti-Inflamatórios/química , Sítios de Ligação , Antígeno CD11c/metabolismo , Cefazolina/química , Proliferação de Células/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Humanos , Interferon gama/metabolismo , Subunidade gama Comum de Receptores de Interleucina/química , Subunidade gama Comum de Receptores de Interleucina/metabolismo , Interleucina-15/metabolismo , Interleucina-2/metabolismo , Janus Quinase 3/metabolismo , Masculino , Monócitos/patologia , Fosforilação/efeitos dos fármacos , Fator de Necrose Tumoral alfa/biossíntese
13.
Immunity ; 52(1): 151-166.e6, 2020 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-31924474

RESUMO

In addition to helper and regulatory potential, CD4+ T cells also acquire cytotoxic activity marked by granzyme B (GzmB) expression and the ability to promote rejection of established tumors. Here, we examined the molecular and cellular mechanisms underpinning the differentiation of cytotoxic CD4+ T cells following immunotherapy. CD4+ transfer into lymphodepleted animals or regulatory T (Treg) cell depletion promoted GzmB expression by tumor-infiltrating CD4+, and this was prevented by interleukin-2 (IL-2) neutralization. Transcriptional analysis revealed a polyfunctional helper and cytotoxic phenotype characterized by the expression of the transcription factors T-bet and Blimp-1. While T-bet ablation restricted interferon-γ (IFN-γ) production, loss of Blimp-1 prevented GzmB expression in response to IL-2, suggesting two independent programs required for polyfunctionality of tumor-reactive CD4+ T cells. Our findings underscore the role of Treg cells, IL-2, and Blimp-1 in controlling the differentiation of cytotoxic CD4+ T cells and offer a pathway to enhancement of anti-tumor activity through their manipulation.


Assuntos
Granzimas/imunologia , Neoplasias/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Proteínas com Domínio T/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/transplante , Transferência Adotiva , Animais , Linhagem Celular Tumoral , Humanos , Interferon gama/imunologia , Interleucina-2/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linfócitos T Reguladores/citologia , Microambiente Tumoral/imunologia
14.
J Immunol ; 204(4): 914-922, 2020 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-31924650

RESUMO

Tick-borne encephalitis (TBE) is a viral infection of the CNS caused by TBE virus. With no specific treatment available, the only protection is a formalin-inactivated whole virus vaccine. Primary immunization with European TBE vaccines, as recommended by the manufacturers, consists of three vaccine doses administered within a 1-y period. Protection from vaccination is believed to be mediated by Abs, yet T cells may also have a protective role. We set out to characterize the human CD4+ T cell response throughout primary TBE immunization. The responses were evaluated before vaccination and 1 mo after each vaccine dose. A heterogeneous magnitude of CD4+ T cell-mediated memory responses was observed in regard to lymphoblast expansion and cytokine production (IFN-γ, IL-2, and TNF), with the highest median magnitude detected after the second dose of vaccine. Stimulation with an overlapping peptide library based on structural TBE virus proteins E and C revealed that CD4+ T cells concomitantly producing IL-2 and TNF dominated the responses from vaccinees after each vaccine dose, whereas a control cohort of TBE patients responded mainly with all three cytokines. CD107a expression was not upregulated upon peptide stimulation in the vaccinees. However, CD154 (CD40L) expression on cytokine-positive memory CD4+ T cells significantly increased after the second vaccine dose. Taken together, TBE vaccination induced CD4+ T cell responses dominated by IL-2 and TNF production together with CD154 upregulation and a lower IFN-γ response compared with TBE patients. This response pattern was consistent after all three doses of TBE vaccine.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Vírus da Encefalite Transmitidos por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/prevenção & controle , Imunogenicidade da Vacina , Memória Imunológica , Vacinas Virais/imunologia , Adulto , Linfócitos T CD4-Positivos/metabolismo , Ligante de CD40/imunologia , Ligante de CD40/metabolismo , Estudos de Casos e Controles , Encefalite Transmitida por Carrapatos/sangue , Encefalite Transmitida por Carrapatos/imunologia , Encefalite Transmitida por Carrapatos/virologia , Feminino , Humanos , Esquemas de Imunização , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-2/imunologia , Interleucina-2/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima/imunologia , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Vacinas Virais/administração & dosagem , Adulto Jovem
15.
Chem Pharm Bull (Tokyo) ; 68(1): 91-95, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31902904

RESUMO

Magnolia Flower is a crude drug used for the treatment of headaches, toothaches, and nasal congestion. Here, we focused on Magnolia kobus, one of the botanical origins of Magnolia Flower, and collected the flower parts at different growth stages to compare chemical compositions and investigate potential inhibitory activities against interleukin-2 (IL-2) production in murine splenic T cells. After determining the structures, we examined the inhibitory effects of the constituents of the bud, the medicinal part of the crude drug, against IL-2 production. We first extracted the flower parts of M. kobus from the bud to fallen bloom stages and analysed the chemical compositions to identify the constituents characteristic to the buds. We found that the inhibitory activity of the buds against IL-2 production was more potent than that of the blooms. We isolated two known compounds, tiliroside (1) and syringin (2), characteristic to the buds from the methanol (MeOH) extract of Magnolia Flower. Moreover, we examined the inhibitory activities of both compounds against IL-2 production and found that tiliroside (1) but not syringin (2), showed strong inhibitory activity against IL-2 production and inhibited its mRNA expression. Thus, our strategy to examine the relationship between chemical compositions and biological activities during plant maturation could not only contribute to the scientific evaluation of medicinal parts of crude drugs but also assist in identifying biologically active constituents that have not yet been reported.


Assuntos
Interleucina-2/metabolismo , Magnolia/química , Extratos Vegetais/química , Animais , Linhagem Celular , Flavonoides/química , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Flores/química , Flores/metabolismo , Glucosídeos/química , Glucosídeos/isolamento & purificação , Glucosídeos/farmacologia , Interleucina-2/genética , Magnolia/metabolismo , Camundongos , Fenilpropionatos/química , Fenilpropionatos/isolamento & purificação , Fenilpropionatos/farmacologia , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo
16.
Drug Dev Ind Pharm ; 46(2): 219-226, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31976759

RESUMO

High consumption of oil formulations has been reported to reduce the blood exposure of drugs like tacrolimus. Consumption of oil formulations has also been shown to inhibit T-cell production of interleukin-2 (IL-2) compared to solid dispersion formulations (SDFs). However, a large amount of oil causes gastrointestinal side effects such as diarrhea and low compliance. Here, we investigated the feasibility of reducing the amount of oil and substitution of chemically synthetized oils for natural oils in these formulations. Reducing the amount of sunflower oil increased blood tacrolimus exposure despite sufficient suppression of IL-2 production. While medium-chain triglyceride (MCT) increased tacrolimus blood exposure, addition of 10% glyceryl monostearate (GMS) to MCT significantly decreased drug blood exposure without requiring a large amount of oil (p < .05). Effects of the contents of GMS in the MCT/GMS formulations, and fatty acid composition in GMS on drug blood exposure were also investigated. The results indicated that both the amount and type of oil were important for maintaining a good balance between a reduction in blood exposure and sufficient IL-2 suppression. The ratio of drug concentration in lymphocytes to that in whole blood after dosing with an oil formulation was significantly higher than that after administration of the SDF (p < .01). These results indicate the feasibility of developing oral oil tacrolimus formulations to reduce systemic side effects and maintain high efficacy for practical use in patients.


Assuntos
Linfócitos/efeitos dos fármacos , Óleos/química , Tacrolimo/administração & dosagem , Tacrolimo/química , Animais , Química Farmacêutica/métodos , Alimentos/efeitos adversos , Glicerídeos/química , Interleucina-2/metabolismo , Linfócitos/metabolismo , Masculino , Ratos , Ratos Endogâmicos Lew , Óleo de Girassol/química , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Triglicerídeos/química
17.
Proc Natl Acad Sci U S A ; 117(1): 285-291, 2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31871161

RESUMO

The impact of ultrasmall nanoparticles (<10-nm diameter) on the immune system is poorly understood. Recently, ultrasmall silica nanoparticles (USSN), which have gained increasing attention for therapeutic applications, were shown to stimulate T lymphocytes directly and at relatively low-exposure doses. Delineating underlying mechanisms and associated cell signaling will hasten therapeutic translation and is reported herein. Using competitive binding assays and molecular modeling, we established that the T cell receptor (TCR):CD3 complex is required for USSN-induced T cell activation, and that direct receptor complex-particle interactions are permitted both sterically and electrostatically. Activation is not limited to αß TCR-bearing T cells since those with γδ TCR showed similar responses, implying that USSN mediate their effect by binding to extracellular domains of the flanking CD3 regions of the TCR complex. We confirmed that USSN initiated the signaling pathway immediately downstream of the TCR with rapid phosphorylation of both ζ-chain-associated protein 70 and linker for activation of T cells protein. However, T cell proliferation or IL-2 secretion were only triggered by USSN when costimulatory anti-CD28 or phorbate esters were present, demonstrating that the specific impact of USSN is in initiation of the primary, nuclear factor of activated T cells-pathway signaling from the TCR complex. Hence, we have established that USSN are partial agonists for the TCR complex because of induction of the primary T cell activation signal. Their ability to bind the TCR complex rapidly, and then to dissolve into benign orthosilicic acid, makes them an appealing option for therapies targeted at transient TCR:CD3 receptor binding.


Assuntos
Ativação Linfocitária/efeitos dos fármacos , Nanopartículas/química , Complexo Receptor-CD3 de Antígeno de Linfócitos T/efeitos dos fármacos , Complexo Receptor-CD3 de Antígeno de Linfócitos T/metabolismo , Dióxido de Silício/química , Dióxido de Silício/farmacologia , Antígenos CD28/metabolismo , Complexo CD3/química , Complexo CD3/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Interleucina-2/metabolismo , Modelos Moleculares , Fosforilação , Complexo Receptor-CD3 de Antígeno de Linfócitos T/química , Complexo Receptor-CD3 de Antígeno de Linfócitos T/genética , Transdução de Sinais/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismo
18.
Immunol Cell Biol ; 98(2): 152-164, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31845380

RESUMO

Depending on the microenvironment conditions, macrophages display phenotypic and functional heterogeneity. This study characterized the programmed cell death-ligand 2 (PD-L2)-expressing macrophage-like cells drained from surgical wound zones, and investigated their influence on helper T (Th) cell responses. Although all CD14+ myeloid cells possessed macrophage-like features, CD206+ and CD163+ cells constituted a specific subpopulation with high PD-L2 expression. There was a modest correlation between the PD-L2 levels on CD206+ macrophages and the amount of interferon (IFN)-γ in the drainage fluid. The adhesion-independent macrophages simultaneously presented both classically-activated M1 and alternatively-activated M2 characteristics. CD206+ and PD-L2+ cells were identified with high granularity and size, expressed arginase-1 and costimulatory molecules, had enhanced phagocytic activity and produced reactive oxygen species. The genes associated with macrophage differentiation (MERTK, AXL and TYRO3) were also upregulated. These cells provided costimulation to Th cells; yet, when PD-L2 was blocked, T-cell proliferation and IFNγ production were enhanced. Under defined conditions devoid of activation stimuli and matrix adhesion, ex vivo-generated monocyte-derived macrophages displayed limited capacity to stimulate T cells. Upon exposure to IFNγ, they significantly upregulated programmed death 1 ligands, especially PD-L2. These cells did not completely abrogate T-cell differentiation; however, PD-L2 checkpoint blockade restored Th1 proliferation and secretion of interleukin-2, tumor necrosis factor-α and IFNγ. In conclusion, upregulation of PD-L2 on the wound zone macrophages may constitute a negative feedback loop that restrains the Th1 effector responses and avoids exacerbation of inflammation during tissue healing.


Assuntos
Diferenciação Celular/imunologia , Inflamação/metabolismo , Macrófagos/metabolismo , Proteína 2 Ligante de Morte Celular Programada 1/metabolismo , Células Th1/imunologia , Cicatrização/fisiologia , Adulto , Apoptose/efeitos dos fármacos , Apoptose/imunologia , Arginase/metabolismo , Antígeno B7-H1/metabolismo , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Interferon gama/farmacologia , Interleucina-2/metabolismo , Lectinas Tipo C/metabolismo , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Lectinas de Ligação a Manose/metabolismo , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Receptores de Superfície Celular/metabolismo , Células Th1/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Cicatrização/efeitos dos fármacos , c-Mer Tirosina Quinase/genética , c-Mer Tirosina Quinase/metabolismo
19.
JCI Insight ; 5(2)2020 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-31855578

RESUMO

Anti-programmed cell death protein 1 (anti-PD-1) therapy has become an immunotherapeutic backbone for treating many cancer types. Although many studies have aimed to characterize the immune response to anti-PD-1 therapy in the tumor and in the peripheral blood, relatively less is known about the changes in the tumor-draining lymph nodes (TDLNs). TDLNs are primary sites of tumor antigen exposure that are critical to both regulation and cross-priming of the antitumor immune response. We used multipanel mass cytometry to obtain a high-parameter proteomic (39 total unique markers) immune profile of the TDLNs in a well-studied PD-1-responsive, immunocompetent mouse model. Based on combined hierarchal gating and unsupervised clustering analyses, we found that anti-PD-1 therapy enhances remodeling of both B and T cell compartments toward memory phenotypes. Functionally, expression of checkpoint markers was increased in conjunction with production of IFN-γ, TNF-α, or IL-2 in key cell types, including B and T cell subtypes, and rarer subsets, such as Tregs and NKT cells. A deeper profiling of the immunologic changes that occur in the TDLN milieu during effective anti-PD-1 therapy may lead to the discovery of novel biomarkers for monitoring response and provide key insights toward developing combination immunotherapeutic strategies.


Assuntos
Anticorpos Antineoplásicos/imunologia , Anticorpos Antineoplásicos/farmacologia , Linfócitos B/imunologia , Linfonodos/efeitos dos fármacos , Neoplasias/imunologia , Receptor de Morte Celular Programada 1/efeitos dos fármacos , Linfócitos T/imunologia , Animais , Anticorpos Antineoplásicos/uso terapêutico , Feminino , Interferon gama/metabolismo , Interleucina-2/metabolismo , Linfonodos/imunologia , Linfonodos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Células T Matadoras Naturais , Neoplasias/tratamento farmacológico , Proteômica , Fator de Necrose Tumoral alfa/metabolismo
20.
BMC Pulm Med ; 19(1): 249, 2019 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-31852476

RESUMO

BACKGROUND: The diagnostic criteria for asthma-chronic obstructive pulmonary disease overlap have not been unified. Different studies have used different criteria, and this has led to diagnostic inconsistencies. METHODS: We collected data of patients who were older than 40 years and hospitalised because of chronic bronchial diseases. One hundred and seventy-one patients were included in this study. We compared seven different diagnostic criteria, examined their consistency, and analysed differences among groups classified with each set. RESULTS: The prevalence of ACO ranged between 7.02 and 27.49% depending on the criteria applied. The patients who met the Soler-Cataluna et al. criteria also met the GesEPOC criteria. Rhee has proposed the strictest diagnostic criteria; hence, the number of patients who met these criteria was the smallest, and those patients also met the diagnostic criteria proposed by the other studies. We found that applying the different sets of criteria did not lead to the selection of the same population, while there were no statistical differences in age, disease duration, allergens, and inflammatory markers. CONCLUSIONS: The diagnostic criteria of ACO have not been unified, which hinders the design and progress of clinical studies that would investigate the ACO phenotypes and underlying mechanisms.


Assuntos
Síndrome de Sobreposição da Doença Pulmonar Obstrutiva Crônica e Asma/diagnóstico , Idoso , Síndrome de Sobreposição da Doença Pulmonar Obstrutiva Crônica e Asma/epidemiologia , Síndrome de Sobreposição da Doença Pulmonar Obstrutiva Crônica e Asma/metabolismo , Síndrome de Sobreposição da Doença Pulmonar Obstrutiva Crônica e Asma/fisiopatologia , Testes Respiratórios , China/epidemiologia , Feminino , Volume Expiratório Forçado , Humanos , Interleucina-2/metabolismo , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Masculino , Pessoa de Meia-Idade , Óxido Nítrico/metabolismo , Prevalência , Espirometria , Fator de Necrose Tumoral alfa/metabolismo
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